Formate dehydrogenase (FDH, EC 1.2.1.2) catalyzes formic acid to generate carbon dioxide and generates NADH along with the reduction of NAD+. Due to various advantages that the reaction is an irreversible reaction, a substrate is cheap formic acid and a product is CO2 and the like, the formate dehydrogenase is used as a coenzyme cycling system to be usually combined with other oxidordeuctase to be used for the bioconversion production of important optical active compounds such as hydroxy acid, chiral alcohol, amino acid and the like. Wild-type FDH (CboFDH) sourced from Candida boidinii is used for regenerating NADH in the production of L-tertiary leucine, which is an example of a maximum application scale of the formate dehydrogenase in the industrialized production at present. Along with the precise parsing of a CboFDH three-dimensional structure, it is possible to reasonably design the CboFDH three-dimensional structure and precisely modify molecules.
The wild-type CboFDH has the disadvantages of low specific enzyme activity and poor operation stability, so that it is of great significance for the chiral compound biosynthesis industry to modify the CboFDH in a site-specific mutagenesis manner, improve the specific enzyme activity and stability and increase the efficiency for regenerating the coenzyme NADH by the CboFDH.